2 AFFINITY STATES OF RI ADENOSINE RECEPTORS IN BRAIN MEMBRANES - ANALYSIS OF GUANINE-NUCLEOTIDE AND TEMPERATURE EFFECTS ON RADIOLIGAND BINDING

Abstract

The binding of agonists and antagonists [(-)-N6-phenylisopropyladenosine, 1,3-diethyl-8-phenylxanthine, Gpp(NH)p, 3-isobutylmethyl xanthine, N6-cyclohexyladenosine, theophylline and 2-chloroadenosine] to Ri adenosine receptors of synaptosomal membranes from rat and bovine brain was studied. The effects of guanine nucleotides and temperature were analyzed with the aid of computerized curve fitting. Evidence is presented for 2 different states of the receptor (one of high and one of low affinity for agonists). Antagonists bind to both states with the same affinity. The 2 states are characterized by saturation, competition and kinetic experiments with very similar results. Guanine nucleotides cause transition of the high- to the low-affinity state. The ratio of the Kd values for the 2 affinity states is 90-150 in rat brain but only 10 in bovine brain. The proportions of the 2 affinity states are the same for all agonists tested; in the absence of exogenous guanine nucleotides, 75% of the total receptor populations is in the high-affinity state, whereas in the presence of guanine nucleotides only 5% remain in the high-affinity state. Binding of antagonists to the receptor is enthalpy-driven whereas binding of the agonist (-)-N6-phenylisopropyladenosine to the high-affinity state of the receptor is entropy-driven. Binding of the agonist to the low-affinity state is enthalpy-driven and thus similar to the binding of antagonists. Guanine nucleotides evidently convert the Ri adenosine receptor from a high- to a low-agonist affinity state and agonist binding shows thermodynamic differences from antagonist binding only when it is to the high-affinity state of the receptor.